AMPK and Mechanisms of Glucose Toxicity

描述(申请人提供)：在糖尿病患者和实验动物中，持续的高血糖会导致肝脏和肌肉中的胰岛素抵抗。我们在多个模型中获得的数据表明，这种葡萄糖诱导的胰岛素抵抗与AMP激活的蛋白激酶(AMPK)/丙二酰辅酶A燃料传感和信号网络的失调(AMPK活性降低和/或丙二酰辅酶A浓度增加)有关。拟议的研究将在其中两个模型中验证这一假设，即暴露在高环境葡萄糖浓度下的培养肝细胞(目标1)和葡萄糖灌输的大鼠(目标3)，在这两个模型中，我们都观察到了AMPK和丙二酰辅酶A的前述变化，以及被研究的胰岛素激活Akt的能力受损。此外，我们将尝试开发一个基于细胞的系统，用于在肌肉中使用C2C12细胞来验证这一假说(目标2)。我们将在每个模型中确定AMPK的变化如何在时间上与受损的胰岛素信号(Akt，IRS-PY)、脂代谢产物(丙二酰辅酶A、DAG、LCCoA)的变化以及可能的下游致病事件(如PKC、IKKB-NFB激活)相关。此外，使用RNAi沉默、病毒构建和/或药理学试剂作为工具，我们将确定AMPK和丙二酰辅酶A的变化是否起到因果作用。最后，我们将探讨葡萄糖输注大鼠AMPK活性降低的可能机制。这些研究将对AMPK/丙二酰辅酶A网络的失调既是葡萄糖诱导的胰岛素抵抗的原因又是其治疗目标的假设进行严格的检验。它们还将为理解胰岛素抵抗的发病机制和治疗提供一个潜在的新框架，胰岛素抵抗是一个在2型糖尿病、早发冠心病、NAFLD/NASH和其他与代谢综合征相关的疾病之前就存在的问题。因此，它们可能会对公共卫生产生重要影响。

DESCRIPTION (provided by applicant): In patients with diabetes and experimental animals sustained hyperglycemia leads to insulin resistance in both liver and muscle. Data obtained by us in a number of models suggest that such glucose-induced insulin resistance is related to dysregulation of the AMP-activated protein kinase (AMPK)/malonyl CoA fuel sensing and signaling network (diminished AMPK activity and/or an increase in malonyl CoA concentration). The proposed studies will test this hypothesis in two of these models, cultured hepatocytes exposed to a high ambient glucose concentration (Aim 1) and glucose-infused rats (Aim 3), in both of which we have observed the aforementioned changes in AMPK and malonyl CoA, and, where studied, an impaired ability of insulin to activate Akt. In addition, we will attempt to develop a cell-based system for testing this hypothesis in muscle using C2C12 cells (Aim 2). We will determine in each of these models how changes in AMPK relate temporally to impaired insulin signaling (Akt, IRS-PY), alterations in lipid metabolites (malonyl CoA, DAG, LCCoA) and putative downstream pathogenetic events (e.g., PKC, IKKB-NFB activation). In addition, using RNAi silencing, viral constructs and/or pharmacological agents as tools, we will determine whether the changes in AMPK and malonyl CoA play a causal role. Finally, we will explore possible mechanisms for the decrease in AMPK activity in the glucose-infused rats. These studies will provide a rigorous test of the hypothesis that dysregulation of the AMPK/malonyl CoA network can be both a cause of glucose-induced insulin resistance and a target for its therapy. They will also provide a potentially novel framework for understanding the pathogenesis and treatment of insulin resistance, a problem that antedates type 2 diabetes, premature coronary heart disease, NAFLD/NASH and other disorders associated with the metabolic syndrome. Thus, they could have an important impact on public health.